This application centers on suppressing Staphylococcus aureus virulence by tuning the function of novel transcriptional factors with small molecules. Staphylococcus aureus is a human pathogen responsible for most wound and hospital-acquired infections. The extensive use of antibiotics to treat S. aureus infections has led to the emergence of high-level resistances in various strains. Virulence suppression provides an alternative strategy to effectively reduce pathogenic potential without asserting selective pressure for developing resistances. A recent breakthrough in my laboratory has identified the MgrA protein as a key virulence regulator in S. aureus. This protein belongs to the MarR family of transcriptional regulators that controls antibiotic resistance and virulence in various bacteria. We demonstrated that the mgrA knockout strain shows a 10,000-fold reduction of virulence in vivo. Subsequently, we discovered that oxidative stress leads to dissociation of MgrA from its promoter DNA. This discovery is significant in that the host immune response to S. aureus infection is to produce reactive oxygen and nitrogen species to counter the pathogen. The microorganism apparently uses MgrA to sense the oxidative stress generated by the host and regulate a global defensive response. In this application, we aim to fully elucidate the mechanism of MgrA and its regulation pathways, and we propose several strategies to suppress S. aureus virulence by tuning MgrA's function with small molecules. In addition, we propose that two other transcriptional regulators in S. aureus, MgrH1 and SarA, use the same mechanism as MgrA to regulate virulence and defensive pathways. The detailed regulation mechanism of these two proteins, the pathways they control, and modulation of their function with small molecules will be investigated as well. Since MgrA is a key virulence determinant in S. aureus, the proposed work could lead to a new strategy for treating infections, while the concept and principles established here could be applied broadly to study other pathogens.